人轮状病毒NSP4基因变异与功能关系的初步研究

A Preliminary Studies on Relationship Between Gene Variation and Function of NSP4 of Human Rotaviruses

在比较我国人A组轮状病毒一般腹泻患者分离株和重症患者分离株非结构蛋白 (NSP) 4cDNA序列时发现 ,两者在可能与致病性有关的区域 (aa131～ 14 6 )内存在着显著的差异。为进一步探讨这种变异是否与毒力改变有关 ,利用杆状病毒表达载体在昆虫细胞Sf9中表达两种毒株的NSP4 ,通过激光扫描共聚焦显微镜初步观察了它对细胞内钙离子浓度的影响。结果表明 :两种来源的NSP4均可使细胞内钙离子浓度明显升高 ,在 4 8h时大致升高3.1～ 3.4倍 ,96h时升高 5 .6～ 5 .8倍 ,但两种毒株之间的差别并不明显。研究证实 ,人轮状病毒NSP4与以往报道的动物轮状病毒NSP4一样 ,可以引起细胞内钙离子增高 ,即可能与病毒的致病性有关。但重症腹泻毒株SZ1NSP4第 131～ 14 6位氨基酸位点出现的变异并未提高其毒力。轮状病毒的毒力改变可能与其它因素有关。

A comparative study was carried out between the NSP4 cDNA derived from the rotavirus strains isolated from infants with light or heavy clinical symptoms.We found that there were remarkable variations in the presumed 'virulent'region of aa 131-146 as reported in the literatures by others.Two NSP4 (from'light symptom'strain B53 and 'heavy symptom'strain SZ1) were endogenously expressed in recombinant baculovirus infected Sf9 cells to see whether there was any influence upon intracellular metabolism of calcium by using confocal microscopy.The results demonstrated a parallel calcium increase with 3.1 to 3.4 times at 48h,and 5.6 to 5.8 times at 96h post-infection, but without noticeable difference.It is worthwhile to notify the fact that the above original observation had identified the calcium increasing ability of the NSP4 of the human rotavirus. What is more,the variation we observed in the 'heavy symptom'strain SZ1 seemed not to elevate its virulence. The increased virulence of the heavy symptom strain seemed possibly irrelevant to variation of NSP4.